Peptide Set of ECM-Related Proteins in Spinal Cords of an Animal Model of Neuropathic Pain Treated with Spinal Cord Stimulation (DTM and Low-Rate)

This is a proteomics set consisting of peptides identified to proteins (using Uniprot) that are related to extracellular matrix proteins. The objective of the experiment was to evaluate the effect of two spinal cord stimulation (DTM SCS and LR SCS) treatments on the ECM-related protein expression levels in the ipsilateral dorsal quadrant of stimulated spinal cord (L1-L2 vertebral level) of rats that were subjected to the spared nerve injury (SNI) model of neuropathic pain. Proteomics were performed by a thrid party (Cell Signaling Techology, Danvers, MA, USA) using their standard processes. They reported the normalized abundance for every peptide that was matched with a corresponsing protein. Data from four experimental groups were compared. The two SCS treatments groups (DTM and LR), a treatment sham group (No-SCS) and a group of uninjured animals (No-SNI). Protein from 4 specimens per group was extracted from tissues and tripsinized. Triptic peptides of samples in every group were pooled and tagged using a tandem mass tag (TMT) system 10-plex system. They were run in parallel with other samples not relevant to this study. A tagged-peptide mix was used in the spectral runs for internal control. Combined tagged peptides were combined and fractionated via reverse column liquid chromatography (LC) using a linear gradient of acetonitrile in 0.125% formic acid (280 nL/min for 150 minutes). Combinations of eluted fractions were subjected to LC/Tandem Mass Spectrometry (TMS) using multi-notch MS3 methodology. Analyses were conducted using optimized protocols (Cell Signaling Technology, Danvers, MA). Mass spectra were evaluated using SEQUEST and the Core platform from Harvard University, and the Uniprot (uniprot.org) rat protein database was used to search for proteins . Results from the search were filtered on precursor ions (±5 ppm mass accuracy) and further filtered to a 1% protein level false discovery rate (FDR). The data set contains the gene name, protein short name, protein description and the normalized abundance (signal to noise) for each of the peptide's mass spectral peak. For phosphoproteomics, proteins from the same samples used in the whole proteomics were digested with trypsin and the peptides purified by reversed-phase solid-phase extraction, followed by phospho-enrichment using immobilized metal affinity chromatography (IMAC) with iron-based magnetic beads (PTMScan® Fe-IMAC, Cell Signaling Technology, Danvers MA). Unbound peptides were washed out, and immobilized phosphopeptides eluted with basic pH buffer. Reversed-phase purification was performed to purify peptides prior to LC/TMS analysis of pooled peptides from 3-4 biological samples, conducted as described above for whole proteomics following standard techniques developed at Cell Signaling Technology. Corresponding phosphorylated isoforms with normalized abundances obtained from the mass spectrum are also included in a separate file.